Enzymes are proteinaceous in nature and act as biocatalysts to regulate many and varied chemical reactions. They are used in many analytical, medical and industrial applications. For example, enzymes find use in medical applications in dialysis systems to provide an artificial replacement for lost kidney function due to renal failure. The enzyme urease is used to convert urea to ammonia and carbon dioxide in hemodialysis and peritoneal dialysis systems. Many applications of enzyme functionality require the use of enzymes in aqueous systems at specific, fixed locations. A number of enzyme immobilization techniques have been employed to accomplish this including physical entrapment or adsorption, chemical adsorption, electrostatic adsorption, hydrogen bonding, covalent bonding, crosslinking, and encapsulation or microencapsulation. These techniques have not, however, made a dry, solid, enzyme crosslinked product that is stable as a dry, solid composition. Further complicating these efforts is the highly sensitive structure-function relationship which is generally unstable in many enzymes, and therefore subject to disruption through changes in temperature and pH, hydrolysis, and chemical or physical modification. These disruptions typically result in deactivation, or a diminished biocatalytic capacity of the enzyme.
Urease is an enzyme used in kidney dialysis treatment systems. Urease is used, for example, in sorbent dialysis to convert the urea in dialysate into ammonium and bicarbonate. These by-products, and others, can then be removed by sorbent materials, such as activated carbon and ion-exchange materials, so that fresh dialysate can be regenerated. Because this type of dialysis system involves flowing water through a layer of water-soluble enzyme, the immobilization of urease is important for at least the following reasons: (i) if not immobilized, the enzyme can dissolve into the flowing water and be transported throughout the system, effectively rinsing it away from its desired location and rendering the dialysate treatment system useless in a very short period of time; (ii) at the same time, the dissolved enzyme can be transported back to the patient by diffusion into the patient fluid, at which point, any urea in the patient would be converted into ammonium inside the patient.
The stability of urease with respect to its activity, or its capacity to catalyze the hydrolysis of urea, is often the source of failure for covalently immobilized, or crosslinked, urease materials. This instability translates to a short shelf-life making most crosslinked urease materials unfeasible for use in consumer products. Attempts aimed at improving the stability of the enzyme in the crosslinked state have included freeze drying, storage in liquid buffers, storage at low temperatures, or some combination of all of these methods. In addition to very limited success, the cost and logistical complications associated with each of these methods often reduces the employment of crosslinked urease to academic exercises.
There have been cost implications in the past with maintaining the stability of crosslinked urease materials. In addition to this, there can be a high cost prohibitive expense of using purified urease as the enzyme source. Almost all methods and materials involving crosslinked urease employ a purified form of the enzyme. Aside from the limited amount of success achieved with these materials, the expense associated with purifying urease poses a major obstacle in the development of consumer products on a wide scale.
A need exists for immobilized enzymes that are stably bound and not capable of dissolution or displacement into a liquid phase. A further need exists for immobilized enzymes that possess a high level of enzymatic activity and maintain steady activity when stored for long periods under ambient conditions. Another need exists for immobilized enzymes that can be produced from economically feasible sources.